Inhibitor-resistant OXY-2-derived β-lactamase produced by Klebsiella oxytoca

Abstract

Klebsiella oxytoca strains are generally moderately resistant to amoxicillin and ticarcillin due to the activities of the chromosomally encoded OXY-1 and OXY-2 class A β-lactamase families. These enzymes have the ability to hydrolyze not only penicillins but also cephalosporins, including cefuroxime, ceftriaxone, and aztreonam, and are inhibited by clavulanic acid. A Klebsiella oxytoca strain was isolated from a culture of blood from a patient who had been treated with amoxicillin-clavulanate (3 g/day) for 10 days 1 month earlier. This strain harbored an unusual phenotype characterized by resistance to amoxicillin-clavulanate. It produced an OXY-2-type β- lactamase (pI 6.3), as confirmed by PCR amplification with primers specific for the OXY-2-encoding gene. Gene sequencing revealed a point mutation (A→G) corresponding to the amino acid substitution Ser→Gly at position 130. This mutant enzyme was poorly inhibited by inhibitors, and its kinetic constants compared to those of the parent enzyme were characterized by an increased K(m) value for ticarcillin, with a drastically reduced activity against cephalosporins, as is observed with inhibitor-resistant TEM enzymes. The substitution Ser→Gly-130 was previously described in the inhibitor-resistant β-lactamase SHV-10 derived from an SHV-5 variant, but this is the first report of such a mutant in OXY enzymes from K. oxytoca.